3 replies to this topic

[HOYAJM]

I have been having problems with degradation of some sort during my protein expression/purification. The same contaminating bands are present and using advice from my previous post here I would like to try a protease inhibitor cocktail. I have small amounts of multiple inhibitors but I am looking for advice on what concentrations to use. I will be adding the inhibitors to 20ml of lysis buffer to resuspend the pellet. What working concentrations are used?

The drugs I have are:

AEBSF
Bestatin
Pepstatin A
E-64

I am thinking about using Roche cOmplete inhibitors but they do not list the concentrations of the inhibitors. Has anyone made their own cocktail and used it successfully to eliminate contaminating bands?

Thanks

[mdfenko]

here's a technical bulletin from sigma which gives information on preparing stocks, range of working concentrations, etc.

Attached Files

•  protease inhibitors-sigma.pdf   87.3K   199 downloads

[HOYAJM]

Just what I was looking for! Thanks mdfenko

[Curtis]

just to add to mdfenko's post, some protease inhibitors (like PMSF) don't disollve in water. you need to dissolve in iso propanol. I used Roche cOmplete. it's good, but it's a little pricey in here.
ps. I can't download this file